The following descriptions and examples are not admitted to be prior art by virtue of their inclusion within this section.
Compartmentalization is a technique that is becoming increasingly popular in the molecular diagnostics and life science research fields. Applications include digital polymerase chain reaction (PCR), two-stage PCR multiplexing (including genotyping), single-cell analysis, targeted sequencing, multiplex immunoassays, ultra-sensitive immunoassays, and library prep for sequencing. Each separate application places different demands on the number of compartments, monodispersity of each compartment, and the volume of each compartment.
One approach for compartmentalizing reactions is by using droplets, which are isolated volumes of a first fluid that are completely surrounded by a second fluid or by a second fluid and one or more surfaces. In the molecular diagnostics and life science research fields this is typically two immiscible liquids. Techniques for droplet generation include co-flow, flow focusing, and T-junction. Co-flow droplet generation forms droplets via pinching of the inner flow from an orifice in a co-flow design as described by, for example, David Weitz (“Monodisperse emulsion generation via drop break off in a coflowing stream,” Langmuir, 2000). Stone and Weitz (“Monodisperse double emulsions generated from a microcapillary device,” Science, 2005) demonstrated double emulsions using a modified co-flowing technique. Flow focusing uses a co-flow design which is geometrically confined in the channel to produce droplets (see, e.g., Stone, “Formation of dispersions using “flow focusing” in microchannels,” APL, 2003). T-junction droplet generation methods and modifications thereof (e.g., Y-junction, cross junction, ψ-junction) generally involve intersecting flows of continuous and dispersed phases (see, e.g., Quake, “Dynamic pattern formation in a vesicle-generating microfluidic device”, PRL, 2001; and Weitz, D. A., Stone, H., “Geometrically mediated breakup of drops in microfluidic devices,” PRL, 2004). Additionally, U.S. Pat. No. 7,943,671 (incorporated herein by reference) described a step emulsification technique that employed an abrupt change in the aspect ratio of a single microchannel to rapidly destabilize a confined co-flowing stream.
The droplet generation techniques described above all require flows of both continuous and dispersed phases. In contrast, Sugiura et al. described a technique in which droplet formation was driven largely by interfacial tension (Sugiura, S., Nakajima, M. “Interfacial tension driven monodispersed droplet formation from microfabricated channel array,” Langmuir, 17:5562-5566 (2001)). With this technique, droplets are generated via falling off a ledge after ejection from a fluidic channel. More recently, Dangla et al., have also described techniques for generating droplets by modulating the interfacial curvature between immiscible liquids using a sloped ceiling to produce a continuously increasing gap height, called a gradient of confinement (U.S. Pat. Pub. 2013/0078164 (incorporated herein by reference); Dangla et al., “Droplet microfluidics driven by gradients of confinement,” PNAS, 10(3):853-858 (2013)). This gradient of confinement has similarities with the interfacial curvature modulation achieved with a discrete step as described by Sugiura et al. (see above).